Supplementary Information

A label-free ratiometric fluorescence nanoprobe for ascorbic acid

based on redox-modulated dual-emission signals

Yuhao Shenga, Tingwei Caoa, Yan Xiaoa,*, Xiuhua Zhanga, Shengfu Wanga, Zhihong Liub

a Hubei Collaborative Innovation Center for Advanced Organic Chemical Materials,

Ministry of Education Key Laboratory for the Synthesis and Application of Organic

Functional Molecules & College of Chemistry and Chemical Engineering, Hubei

University, Wuhan 430062, Chinab Key Laboratory of Analytical Chemistry for Biology and Medicine (Ministry of

Education), College of Chemistry and Molecular Sciences, Wuhan University, Wuhan

430072, China.

Experiment section

Materials and instruments

The polymer poly(9,9-dioctylfluorenyl-2,7-diyl) (PFO, Mw=10000~100000 by

GPC) was purchased from Xi’an Polymer Light Technology Corp. Amphiphilic

functional polymer poly(styrene-co-maleic anhydride) (PSMA, terminated by

cumene, content of 68% styrene, average molecular weight about 1700), L-

glutathione-reduced (GSH), gold (III) chloride trihydrate (HAuCl4·3H2O) and 4-(2-

hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES) were purchased from Sigma-

Aldrich (St. Louis, MO, USA). Tetrahydrofuran (THF, anhydrous, 99.9%) and

sodium hypochlorite pentahydrate (NaClO·5H2O) were purchased from Sinopharm

Chemical Reagent Co., Ltd. (Shanghai, China). Ascorbic acid (AA), metal salt

compounds, amino acid, bovine serum albumin (BSA), trypsin, and glucose were

purchased from Aladdin (Shanghai, China). All of the chemicals were of analytical

grade without further treatment. The glassware used in the experiments were all

rinsed by aqua regia (HCl/HNO3 = 3:1, v/v). Ultrapure water was obtained through

Electronic Supplementary Material (ESI) for Analyst.This journal is © The Royal Society of Chemistry 2019

Merck Millipore water purification system with an electric resistance 18.25 MΩ.

Human serum was provided by Zi Jing Hospital (Wuhan, Hubei).

The fluorescence measurements were performed by RF-5301 PC spectrometer

(Shimadzu, Japan). The UV-vis absorption spectra were measured on a UV-2550 UV-

vis spectrometer (Shimadzu, Japan). TEM images were obtained on JEM-2100

transmission electron microscope with an acceleration voltage of 200 kV (Hitachi,

Japan). Zetasizer Nano (Malvern) was used to measure zeta potential of the

nanomaterials. FT-IR spectrum was conducted on a Magan-IR Spectrometer 500

(Nicolet, Madision, WI, USA) with the KBr pellet technique. X-ray photoelectron

spectroscopy (XPS) spectra were obtained from a Kratos Axis Ultra LD spectrometer

using a monochromatic Al Kα source (PerkinElmer, USA). The time-resolved

photoluminescence were measured by a FLS980 Lifetime Spectrometer (Edinburgh,

England).

Synthesis of poly(9,9-dioctyluorenyl-2,7-diyl) dots (PFO dots)

The PFO dots were synthesized according to the literature.1 Briefly, the polymer

PFO and amphiphilic copolymer PSMA were dissolved in tetrahydrofuran (THF) to

reach a stock solution of 1 mg/mL, respectively. 250 μL of PFO and 100 μL of PSMA

were mixed and diluted with 4.65 mL THF to produce a solution mixture which

contains 50 μg/mL PFO and 20 μg/mL PSMA. The mixture was sonicated to reach a

homogeneous solution. Subsequently, 5 mL of mixture solution was added into 10 mL

of ultrapure water quickly in a bath sonicator. THF was removed by nitrogen stripping

overnight, followed by filtration through a 0.22 μm filter to remove the fraction of

aggregates. The resulting PFO dots dispersions were clear and stable for several

months without signs of aggregation.

Preparation of CoOOH nanoflakes

The fabrication of CoOOH nanoflakes was performed according to the reported

study with minor revision.2 Typically, NaClO (0.2 M, 10 mL) and NaOH (0.8 M, 10

mL) were added into CoCl2 solution (10 mM, 20 mL) at room temperature, and then

the mixture was sonicated for 10 min. After that, the CoOOH nanoflakes were

collected from the suspension by centrifugation at 8000 rpm for 10 min and the

products were washed several times with ultrapure water to remove the unreacted

substances. The as-prepared CoOOH nanoflakes were redissolved with ultrapure

water and further treated by sonication to obtain homogeneous CoOOH nanoflakes

solution.

Preparation and purification of GSH-AuNCs

GSH-AuNCs were synthesized by chemical reduction of HAuCl4 with

glutathione.3 A solution of glutathione (5 mL, 15 mM) was added to an equal volume

of 10 mM HAuCl4. The mixture was vigorously stirred till the solution slowly turn to

the faint yellow, followed by the addition of 1 mL of 1 M NaOH. After the solution

reaching the colorless with the stirring, the solution was incubated at 37 ˚C under

continuous stirring for 24 h. Finally, the solution was extensively dialyzed against

ultrapure water using a dialysis tube (MWCO = 3500 Da) overnight.

Figures

Figure S1. The EDS spectrum of the CoOOH nanoflakes.

Figure S2. (A) TEM image of PFO dots, the inset image is the size distributions. (B)

UV-vis absorption spectrum (black line) and fluorescence emission spectrum (blue

line) of PFO dots. (C) FT-IR spectra of (a) PFO monomer and (b) PFO dots.

Figure S3. Zeta Potential of (a) PFO dots, (b) CoOOH, (c) GSH-AuNCs, (d) PFO

dots/CoOOH/GSH-AuNCs nanoprobe.

Figure S4. (A) TEM image of GSH-AuNCs. Inset image: the size distributions and

the lattice fringes of GSH-AuNCs (HRTEM). (B) The UV-vis absorption (black line)

and fluorescence emission (blue line) spectra of GSH-AuNCs. (C-D) The XPS spectra

of Au 4f (C) and S 2p (D) for GSH-AuNCs. (E) FT-IR spectra of (a) GSH and (b)

GSH-AuNCs.

Figure S5. UV-vis absorption spectrum of CoOOH nanoflakes (black line) and

fluorescence spectrum of PFO dots (blue line).

Figure S6. (A) Time-resolved fluorescence spectra of PFO dots in the absence (black

line) and presence (red line) of CoOOH nanoflakes. Excitation wavelength was 380

nm. (B) The fluorescence spectra of PFO dots (black line), PFO dots incubated with

20 μg/mL CoOOH (red line) and PFO dots incubated with CoOOH following the

addition of 100 μM AA (blue line).

Table S1. Decay Parameters for PFO dots without and with CoOOH nanoflakesNames τ1/ns τ2/ns τ3/nsPFO dots 0.39 2.98 9.36

17.32% 36.57% 46.10%PFO dots + CoOOH 0.39 2.71 8.54

22.75% 42.77% 34.48%

All data were fitted by a cubic exponential decay function.

Figure S7. The redox reaction between CoOOH and AA.

Figure S8. (A) The fluorescence spectra of AuNCs with different concentration of

Co2+ (0-0.2 μM). (B) The fluorescence spectra of AuNCs (black line), AuNCs + AA

(red line), AuNCs + PFO dots (blue line), AuNCs + CoOOH (pink line) and AuNCs +

CoOOH + AA (green line). (C) UV-vis absorption spectra of AuNCs (black line) and

CoOOH nanoflakes (red line).

Figure S9. (A) Time-resolved fluorescence spectra of the AuNCs in the absence

(black line) and presence (red line) of Co2+. Excitation wavelength was 380 nm. (B)

Emission spectra of the AuNCs in the absence (black line) and presence (red line) of

0.2 μM Co2+, and in the presence of 0.2 μM Co2+ with 0.2 μM EDTA (blue line).

Table S2. Decay Parameters for AuNCs without and with Co2+

All data were fitted by a bi-exponential decay function.

Figure S10. (A) Fluorescence quenching effect of PFO dots by varying amounts (0-

30 μg/mL) of CoOOH nanoflakes. (B) Fluorescence intensity of PFO dots in response

to different incubating time with CoOOH nanoflakes.

Figure S11. TEM image of the as-constructed nanoprobe.

Figure S12. Fluorescence response of the nanoprobe in the absence (red line) and presence (black line) of 100 μM AA as a function of time.

Names τ1/ns τ2/nsAuNCs 49.91 587.68

7.83% 92.17%AuNCs + Co2+ 41.45 546.56

17.79% 82.21%

Table S3. Comparison with other reported methods for AA detection

Method ProbeLinear range

(μM)Detection limit (μM)

References

Amperometry RGO–ZnOa 50-2350 3.71 4

Amperometry CTAB/rGO/ZnSb 50-1000 30 5

Voltammetry PCN-333 (Al) MOFsc 14.1-5500 4.60 6

Voltammetry CL-TiNd 50-1500 1.52 7

DPV Pd@Au/RGOe 50-2856.6 2.88 8

Fluorescence Hydroxycoumarin/MnO2 10-50 1.6 9

Fluorescence Fe3+/Fe2+-CdTe QDs@SiO2 3.33-400 1.25 10

Fluorescence PFO dots/CoOOH/AuNCs 10-300 1.9 This work

a RGO-ZnO-reduced graphene oxide-zinc oxideb CTAB-hexadecyl trimethyl ammonium bromide, ZnS-zinc sulfide nanocompositec Porous coordination network, Metal-organic frameworksd Chrysanthemum-like titanium nitridee Pd@Au-Pd-Au core-shell, RGO-reduced graphene oxide, GCE-glassy carbon electrode.

Figure S13. Viability of Hela cells treated with PFO dots/CoOOH/AuNCs nanoprobe.

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